胶质瘤中MicroRNA-184的表达及其预后价值

目的 研究人脑胶质瘤石蜡包埋组织（formalin-fixed paraffin-embedded, FFPE）中微小RNA-184（microRNA-184, miR-184）的表达及其与临床病理特征之间的关系,探讨其在预测胶质瘤患者预后中的价值。方法 采用微阵列芯片法和RT-PCR检测108例胶质瘤FFPE和32例正常脑组织中的miR-184表达,同时Kaplan-Meier法分析胶质瘤患者总生存期和无病生存期情况。结果 胶质瘤组织中miR-184的表达下调,并用RT-PCR进一步验证; miR-184低表达与WHO分级（P=0.007）、Karnofsky 评分（KPS）（P=0.029）、复发时间（P=0.037）和生存时间（P=0.019）显著相关;Kaplan-Meier 分析结果表明,miR-184低表达与高表达患者总生存期（OS）（P=0.001）和无病生存期（DFS）（P=0.006）差异有统计学意义,miR-184低表达患者预后较差。多因素分析显示胶质瘤中miR-184的差异表达是预测OS[风险比（HR）：7.52;95％CI：2.63~21.42;P=0.002]和DFS[HR：11.56;95％CI：5.17~25.93;P<0.001]的独立危险因素。结论 miR-184的表达与胶质瘤患者预后显著相关,表明miR-184可作为预测胶质瘤患者预后的独立标志物。     

MicroRNA-195 inhibits non-small cell lung cancer cell proliferation,migration and invasion by targeting MYB

MicroRNA-195 (miR-195) has been implicated in several other cancers; however, its role in non-small cell lung cancer (NSCLC) remains unclear. In this study, we demonstrated that miR-195 was significantly down-regulated in NSCLC samples and cell lines compared with corresponding normal counterparts. In vitro and in vivo functional assays demonstrated that modulation of miR-195 expression affected NSCLC cell proliferation, migration and invasion. Using miRNA target prediction algorithms and reporter assays, we demonstrated that miR-195 suppressed the expression of MYB both at the mRNA and protein level, and was directly bound to the 3′untranslated region of MYB mRNA. Overexpression of MYB in NSCLC cells using an ectopic expression vector restored the decreased cell proliferation, migration and invasion effects induced by miR-195. Finally, we observed an inverse correlation between MYB and miR-195 in NSCLC. Taken together, our findings indicated that miR-195 functions as tumour suppressor in NSCLC, and the miR-195/MYB axis might represent a potential therapeutic target for NSCLC intervention.     

microRNA-122抑制胰腺癌细胞增殖、迁移和侵袭

目的 明确microRNA-122(miR-122)在胰腺癌中的表达及miR-122对胰腺癌细胞增殖、迁移和侵袭的作用.方法 采用RT-qPCR方法检测20例胰腺癌组织、胰腺癌细胞株(ASPC1、PANC1、MP和SW1990)和人源正常胰腺导管上皮细胞株(HPDE)中miR-122的表达量.将miR-122模拟类似物...     

miR-203a-3p通过靶向调控GATA6抑制食管鳞癌细胞的增殖和侵袭

背景与目的：生物信息学分析提示GATA6是miR-203a-3p的潜在靶基因,明确miR-203a-3p通过靶向调控GATA6抑制食管鳞癌细胞的增殖和侵袭。方法：采用Lipofectamine ^TM RNAiMAX对培养的KYSE-70和KYSE-180细胞瞬时转染。采用实时荧光定量聚合酶链反应（real-time fluorescence quantitative polymerase chain reaction,RTFQ-PCR）检测miR-203a-3p和GATA6的表达水平。采用蛋白质印迹法（Western blot）检测GATA6蛋白的水平。质粒联合转染后检测相对萤光素酶活性。对食管鳞癌患者标本进行恶性肿瘤与异型增生组织miR-203a-3p和GATA6的表达检测。结果：RTFQ-PCR及Western blot检测结果显示,与对照组比较,GATA6基因和蛋白的表达在miR-203a-3p转染组中降低,在miR-203a-3p inhibitor组却升高,差异均有统计学意义（P<0.05）。与对照组比较,miR-203a-3p转染组KYSE-70细胞增殖能力下降,miR-203a-3p inhibitor组中却升高,差异均有统计学意义（P<0.05）。在KYSE-180中虽然差异无统计学意义,但其趋势却与KYSE-70一致。与对照组比较,在KYSE-70和KYSE-180细胞系中,侵袭细胞数值/视野在miR-203a-3p转染组中均明显下降（P<0.01）,在miR-203a-3p inhibitor组中却明显升高（P<0.05）。与miR-203a-3p掠夺型+GATA6野生型组和miR-203a-3p野生型+GATA6突变型组比较,相对萤光素酶活性在miR-203a-3p野生型+GATA6野生型组中降低,差异有统计学意义（P<0.05）。与异型增生组织相比较,100%（10/10）食管鳞癌患者miR-203a-3p在恶性肿瘤组织的表达下调,而GATA6表达水平上调。结论：miR-203a-3p通过靶向调控GATA6抑制食管鳞癌细胞的增殖和侵袭能力。     

LncRNA WEE2-AS1 knockdown inhibits the proliferation,migration and invasion of glioma cells via regulating miR-29b-2- 5p/TPM3 axis

Glioma is a general malignant tumor with a dismal prognosis. Long noncoding RNAs (lncRNAs) have been implicated in the initiation and processes of tumors. An investigation of the GEPIA database revealed that long noncoding RNA WEE2 antisense RNA 1 (WEE2-AS1) is upregulated in glioma tissues compared to normal brain tissues, and validation with quantitative real-time polymerase chain reaction (qRT–PCR) revealed that WEE2-AS1 expression was consistent with the database prediction. Fluorescence in situ hybridization (FISH) assays revealed that WEE2-AS1 was localized primarily in the cytoplasm. Clone formation experiment and EDU assay were used to detect cell proliferation ability, and Transwell assay was used to detect cell migration and invasion ability, Western-blot assay and immunofluorescence were used to determine TPM3 protein level. Functional experiments revealed that the downregulation of WEE2-AS1 impeded cell proliferation, migration, and invasion in glioma cell lines. Furthermore, downregulation of WEE2-AS1 suppressed tumor growth in vivo. Bioinformatics predictions and integrated experiments indicated that WEE2-AS1 promoted tropomyosin 3 (TPM3) expression by sponging miR-29b-2-5p. A dual-luciferase reporter assay was conducted to uncover the binding of WEE2-AS1 and miR-29b-2-5p and that of miR-29b-2-5p and TPM3. Additionally, a series of rescue assays showed that WEE2-AS1 promotes proliferation, migration, and invasion by targeting miR-29b-2-5p to regulate TPM3 expression. Ultimately, the results of this study indicate that WEE2-AS1 plays an oncogenic role in glioma and may promote further investigations of the diagnostic and prognostic value of WEE2-AS1 in glioma.     

长链非编码RNA LOC101927476抑制卵巢癌细胞侵袭迁移和增殖

目的探讨长链非编码RNA(LncRNA)LOC101927476在卵巢癌细胞中的表达及其对卵巢癌生物学特征的影响。方法选取2018—2019年于中国医学科学院肿瘤医院手术治疗的卵巢癌患者。采用实时荧光定量PCR(real-time PCR)检测卵巢癌细胞系3AO、OVCA429、TOV21G、A2780、SKOV3以及22例卵巢癌原发瘤和转移瘤组织中LncRNA LOC101927476的表达水平, 采用TCGA数据库中卵巢癌转录组测序数据验证LncRNA LOC101927476和GATA4的表达, 采用慢病毒包装体系构建过表达LOC101927476(OE-LOC101927476组)和空载(OE-EV组)慢病毒并转染至3AO细胞。设计单链向导RNA(sgRNA), 利用CRISPR/Cas9构建LncRNA LOC101927476敲除细胞系。采用Transwell法和划痕实验检测LncRNA LOC101927476对卵巢癌细胞侵袭和转移能力的影响。采用细胞计数盒8(CCK-8)法检测细胞的增殖能力。结果 real-time PCR显示, 22例卵巢癌患者中, 20例转移灶中L...     

miR-34a对结肠癌细胞SW480增殖、迁移和侵袭能力的影响及机制探讨

目的 探讨miR-34a对结肠癌细胞株SW480增殖、侵袭和迁移能力的影响及可能的作用机制。方法将miR-34a过表达慢病毒、空病毒载体转染SW480细胞,未做处理细胞作为空白对照组。Real-time PCR检测各组细胞内miR-34a的表达;CCK8法检测细胞增殖能力;划痕实验、Transwell小室实验检测细胞迁移和侵袭能力;Western blot实验检测细胞内E-cadherin和Vimentin蛋白表达。结果 与空病毒载体组及空白对照组相比,转染组细胞中miR-34a的表达增高,且细胞增殖效率、侵袭和迁移能力降低(P＜0.05),miR-34a使E-cadherin蛋白表达增加、Vimentin蛋白表达降低。结论 miR-34a可抑制结肠癌细胞SW480增殖、侵袭和迁移能力,并能影响E-cadherin和Vimentin的表达,miR-34a有望成为干预结肠癌转移和复发的分子靶点。     

MicroRNA-138 suppresses invasion and promotes apoptosis in head and neck squamous cell carcinoma cell lines

Metastasis is a critical event in the progression of head and neck squamous cell carcinoma (HNSCC). To identify microRNAs associated with HNSCC metastasis, six paired HNSCC cell lines with different metastatic potential were examined. Using microarrays, a panel of differentially expressed microRNAs was identified, including reduction of miR-138 in highly metastatic cells. Ectopic transfection of miR-138 suppressed cell invasion and led to cell cycle arrest and apoptosis. Knockdown of miR-138 enhanced cell invasion and suppressed apoptosis. Thus, our results suggested miR-138 acts as a tumor suppresser and may serve as a therapeutic target for HNSCC patients at risk of metastasis.     

MicroRNA-150对上皮性卵巢癌细胞增殖、凋亡和侵袭转移能力的影响

目的 研究microRNA-150(miR-150)在人上皮性卵巢癌细胞中的表达情况,及其对人上皮性卵巢癌细胞增殖、凋亡和侵袭转移能力的影响。方法 通过荧光实时定量PCR(qRT-PCR)检测各处理组细胞中的miR-150表达水平;利用MTT法、流式细胞技术、Transwell法探究miR-150的表达对上皮性卵巢癌细胞增殖、凋亡及侵袭转移能力的影响。结果 与正常卵巢上皮细胞(T29)相比,上皮性卵巢癌细胞(A2780和OVCAR3)中miR-150表达显著降低(P＜0.01);转染miR-150mimic后,A2780和OVCAR3细胞中miR-150水平明显升高(P＜0.01);MTT试验显示,转染培养三天后,miR-150 mimic组细胞OD值(A2780:1.12±0.03;OVCAR3:1.91±0.03)较Blank组(A2780:2.35±0.09;OVCAR3:2.63±0.07)及miR-150 NC组(A2780:2.18±0.07;OVCAR3:2.43±0.11)明显降低(P＜0.01);细胞凋亡检测显示:miR-150 mimic组凋亡率(A2780:16.10±0.58%;OVCAR3:15.16±1.30%)较Blank组(A2780:10.07±0.66%;OVCAR3:3.81±0.24%)及miR-150 NC组(A2780:10.36±1.08%;OVCAR3:4.89±0.07%)明显升高(P＜0.01);Transwell试验显示:miR-150 mimic组穿膜细胞数(A2780:38.67±2.03;OVCAR3:28.67±2.03)较Blank组(A2780:76.30±7.45;OVCAR3:55.67±3.18)及miR-150 NC组(A2780:74.33±5.78;OVCAR3:56.33±3.84)明显减少(P＜0.01)。结论 miR-150在上皮性卵巢癌细胞中表达降低,可能是上皮性卵巢癌增殖、侵袭转移的机制之一;上调miR-150的表达可以抑制上皮性卵巢癌细胞增殖、促进细胞凋亡,并且降低上皮性卵巢癌细胞侵袭转移能力。     

Overexpression of LACTB,a Mitochondrial Protein That Inhibits Proliferation and Invasion in Glioma Cells

LACTB, a mitochondrial protein, was ubiquitously expressed in different mammalian tissues, such as liver, heart, and skeletal muscle. It has been shown that LACTB is downexpressed in breast cancers, and it suppresses the proliferation and promotes the apoptosis of breast cancers. However, its role in the progression and prognosis of glioma remains unknown. In this study, we analyzed the clinicopathological features and outcomes of LACTB expression in 98 glioma patients and investigated the effects of LACTB overexpression on the proliferation, invasion, and angiogenesis of glioma cells in vitro. We observed a significant decrease in LACTB expression in glioma, and downexpression of LACTB is correlated with a poor prognosis of glioma patients. Moreover, Cox regression analysis reveals that the LACTB is an independent prognostic indicator for glioma patients. Overexpression of LACTB could suppress the proliferation, invasion, and angiogenesis of glioma cells. In addition, overexpression of LACTB could inhibit the expression of PCNA, MMP2, MMP9, and VEGF. Taken together, these data indicate that LACTB may serve as a promising therapeutic target for gliomas.     

敲低DCLK1抑制神经胶质瘤细胞增殖、 迁移与侵袭CSCD

目的探讨双肾上腺皮质激素样激酶 1(DCLK1)对胶质瘤细胞增殖、迁移与侵袭的影响及其机制。方法 RT-qPCR、免疫组织化学染色和Western blot检测胶质瘤组织和细胞(U87和A172)中DCLK1 mRNA和蛋白的表达情况;sh-DCLK1和阴性对照(sh-Con)转染U87和A172细胞;MTT和Transwell法分析敲低DCLK1对神经胶质瘤细胞增殖、迁移和侵袭能力的影响;蛋白印迹法检测敲低DCLK1对神经胶质瘤细胞中TGF-β/Smads信号通路相关蛋白TGF-β1、p-Smad2、p-Smad7表达的影响;将已转染sh-DCLK1或sh-Con的U87细胞注射到BALB/c裸鼠颈部皮下,定期测量瘤体体积,收集瘤体并称重。结果胶质瘤组织和细胞中DCLK1 mRNA和蛋白表达较癌旁正常组织和正常神经胶质细胞均显著升高(P<0.001)。与sh-Con组比,sh-DCLK1组的胶质瘤细胞增殖、迁移和侵袭能力及细胞中TGF-β1、p-Smad2和p-Smad7表达显著降低(P<0.01;P<0.001);sh-DCLK1组裸鼠体内的瘤体体积和瘤体质量明显降低(P<0.001)。结论敲低DCLK1能抑制胶质瘤细胞增殖、迁移和侵袭,其机制可能与抑制TGF-β/Smads信号活性有关。     

下调肌动蛋白相关蛋白2/3复合体5对肺鳞癌细胞株SK-MES-1增殖、侵袭和转移能力的影响

背景与目的：肌动蛋白相关蛋白2/3复合体5(actin related protein 2/3 complex subunit 5,ARPC5)参与肌动蛋白的组装,影响细胞的运动能力,在形成癌细胞侵袭性伪足的过程中起到重要作用。本研究通过基因沉默手段,探讨下调ARPC5基因表达对肺鳞癌细胞株SK-MES-1增殖、侵袭和转移能力的影响。方法：应用阳离子脂质体转染法构建ARPC5基因沉默(si-ARPC5)的SK-MES-1肺鳞癌细胞株,qRT-PCR和蛋白质印迹法(Western blot)鉴定转染效率和表达情况。实验设置si-ARPC5实验组、空白对照组(mock组)及阴性对照组(NC组)。采用细胞技术试剂盒(cell couiting kit-8,CCK-8)法测定si-ARPC5对SK-MES-1细胞的增殖的影响;划痕修复实验测定si-ARPC5对SK-MES-1细胞迁移能力的影响;Transwell小室体外侵袭实验测定si-ARPC5对SK-MES-1细胞的侵袭能力的影响。结果：转染si-ARPC5后的SK-MES-1细胞在mRNA和蛋白水平上均下调ARPC5的表达。与mock组及NC组比较,si-ARPC5实验组SK-MES-1细胞增殖能力明显下降(t=7.993,t=8.681,P<0.05)。si-ARPC5实验组划痕修复率(43.32±0.23)%,而mock组和NC组划痕修复率分别为(73.11±0.43)%和(76.58±0.88)%,差异有统计学意义(t=7.348,t=10.614,P<0.05)。si-ARPC5组的SK-MES-1细胞穿膜数为(27±6)个,mock组为(101±11)个,NC组为(92±9)个。si-ARPC5组穿膜细胞数明显少于其他2组,差异有统计学意义(t=10.229,t=8.391,P<0.05)。结论：下调ARPC5基因的表达能够显著抑制肺鳞癌细胞株 SK-MES-1的增殖、侵袭和转移能力。